1. Field of the Invention
The present application relates to the synthesis of API (active pharmaceutical ingredient) grade 6-amino-2-chloro-9-(2′-deoxy-β-D-ribofuranosyl)-purine, otherwise known as cladribine (I), CdA and 2-chloro-2′-deoxyadenosine, that can be used on manufacturing scales.
2. Description of the Related Art
Previously Robins and Robins (Robins, M. J. and Robins, R. K., J. Am. Chem. Soc. 1965, 87, 4934-4940) reported that acid-catalyzed fusion of 1,3,5-tri-O-acety-2-deoxy-D-ribofuranose and 2,6-dichloropurine gave a 65% yield of an anomeric mixture 2,6-dichloro-9-(3′,5′-di-O-acetyl-2′-deoxy-α-,β-D-ribofuranosyl)-purines from which the α-anomer was obtained as a pure crystalline product by fractional crystallization from ethanol in 32% yield and the equivalent β-anomer remained in the mother liquor (see Scheme 1). The β-anomer, which could have been used to synthesize cladribine, wasn't isolated further. The α-anomer was treated with methanolic ammonia which resulted in simultaneous deacetylation and amination to give 6-amino-2-chloro-9-(2′-deoxy-α-D-ribofuranosyl)-purine, which is a diastereomer of cladribine.

Broom et al. (Christensen, L. F., Broom, A. D., Robins, M. J., and Bloch, A., J. Med. Chem. 1972, 15, 735-739) adapted Robins et al.'s method by treating the acetylated mixture (viz., 2,6-dichloro-9-(3′,5′-di-O-acety-2′-deoxy-α,β-D-ribofuranosyl)-purine) with liquid ammonia and reacylating the resulting 2′-deoxy-α- and -β-adenosines with p-toluoyl chloride (see Scheme 2). The desired 2-chloro-9-(3′,5′-di-O-p-toluoyl-2′-deoxy-β-D-ribofuranosyl)-adenine was then separated by chromatography and removal of the p-toluoyl group resulted in cladribine in 9% overall yield based on the fusion of 1,3,5-tri-O-acety-2-deoxy-D-ribofuranose and 2,6-dichloropurine.

To increase the stereoselectivity in favour of the β-anomer, Robins et al. (Robins, R. L. et al., J. Am. Chem. Soc. 1984, 106, 6379-6382, U.S. Pat. No. 4,760,137, EP0173059) provided an improved method in which the sodium salt of 2,6-dichloropurine was coupled with 1-chloro-2-deoxy-3,5-di-O-p-toluoyl-α-D-ribofuranose in acetonitrile (MeCN) to give the protected β-nucleoside in 59% isolated yield, following chromatography and crystallisation, in addition to 13% of the undesired N-7 regioisomer (see Scheme 3). The apparently higher selectivity in this coupling reaction is attributed to it being a direct SN2 displacement of the chloride ion by the purine sodium salt. The protected N-9 2′-deoxy-β-nucleoside was treated with methanolic ammonia at 100° C. to give cladribine in an overall 42% yield. The drawback of this process is that the nucleophilic 7- position nitrogen competes in the SN2 reaction against the nucleophilic 9- position, leading to a mixture of the N-7 and N-9 glycosyl isomers as well as the need for chromatography and crystallisation to obtain the pure desired isomer.

Gerszberg and Alonso (Gerszberg S. and Alonso, D. WO0064918, and US20020052491) also utilised an SN2 approach with 1-chloro-2-deoxy-3,5-di-O-p-toluoyl-α-D-ribofuranose but instead coupled it with the sodium salt of 2-chloroadenine in acetone giving the desired β-anomer of the protected cladribine in 60% yield following crystallisation from ethanol (see Scheme 4). After the deprotection step using ammonia in methanol (MeOH), the β-anomer of cladribine was isolated in an overall 42% yield based on the 1-chlorosugar, and 30% if calculated based on the sodium salt since this was used in a 2.3 molar excess.

To increase the regioselectivity towards glycosylation of the N-9 position, Gupta and Munk recently (Gupta, P. K. and Munk, S. A., US20040039190, WO2004018490 and CA2493724) conducted an SN2 reaction using the anomerically pure α-anomer, 1-chloro-2-deoxy-3,5-di-O-p-toluoyl-α-D-ribofuranose but coupling it with the potassium salt of a 6-heptanoylamido modified purine (see Scheme 5). The bulky alkyl group probably imparted steric hindrance around the N-7 position, resulting in the reported improved regioselectivity. Despite this, following deprotection, the overall yield of cladribine based on the 1-chlorosugar was 43%, showing no large improvement in overall yield on related methods. Moreover 2-chloroadenine required prior acylation with heptanoic anhydride at high temperature (130° C.) in 72% yield, and the coupling required cryogenic cooling (−30° C.) and the use of the strong base potassium hexamethyldisilazide and was followed by column chromatography to purify the product protected cladribine.

More recently Robins et al. (Robins, M. J. et al., J. Org. Chem. 2006, 71, 7773-7779, US20080207891) published a procedure for synthesis of cladribine that purports to achieve almost quantitative yields in the N-9-regioselective glycosylation of 6-(substituted-imidazol-1-yl)-purine sodium salts with 1-chloro-2-deoxy-3,5-di-O-p-toluoyl-α-D-ribofuranose in MeCN/dichloromethane (DCM) mixtures to give small or no detectable amounts of the undesired α-anomer (see Scheme 6). In actuality this was only demonstrated on the multi-milligram to several grams scale, and whilst the actual coupling yield following chromatography of the desired N-9-β-anomer was high (83% to quantitative), the protected 6-(substituted-imidazol-1-yl)-products were obtained in 55% to 76% yield after recrystallisation. Following this, toxic benzyl iodide was used to activate the 6-(imidazole-1-yl) groups which were then subsequently displaced by ammonia at 60-80° C. in methanolic ammonia to give cladribine in 59-70% yield following ion exchange chromatography and multiple crystallisations, or following extraction with DCM and crystallisation. Although high anomeric and regioselective glycosylation was demonstrated the procedure is longer than the prior arts, atom uneconomic and not readily applicable to industrial synthesis of cladribine such as due to the reliance on chromatography and the requirement for a pressure vessel in the substitution of the 6-(substituted-imidazole-1-yl) groups.

Therefore, there is a need for a more direct, less laborious process, which will produce cladribine in good yield and high purity that is applicable to industrial scales.